The goal of this project is to develop and adapt electron spin resonance spectroscopy to study the biochemistry, physiology, and pathology of cells and tissues in order to answer problems of medical and biological importance. To accomplish this goal we are working on approaches to increase the sensitivity and developing cavity design suitable for different problems ranging from microsamples, to cultured cells to whole tissues. By utilizing different frequency microwave sources it is possible to optimize resonator design for each type of biological sample. Initially we assembled an X-band, 9 GHz spectrometer. Various resonators were designed and tested at X-band including several loop gap resonators. In order to accommodate large aqueous samples such as living perfused organs work was started on development of L band and S band spectrometers. S and L band loop gap resonator were designed and built to enable the study of free radical generation in living perfused hearts. Over the past year we have focused on 2 important cardiovascular applications (1) the mechanism of the adriamycin cardiomyopathy (2) the mechanism of ischemic and reperfusion heart damage. We demonstrated that Fe(III) binds to adriamycin and that these complexes cycle to reduce oxygen. This mechanism explains the formation of reduced oxygen and drug radicals which are thought to mediate adriamycin's therapeutic and toxic effects. Free radicals are thought to be generated in the ischemic and post-ischemic reperfused heart and to mediate cellular damage. We developed a direct ESR technique to measure free radical generation in the ischemic and post ischemic heart. This technique was used to quantitate and characterize free radical generation in the post-ischemic heart. In addition the molecular mechanisms of free radical generation were studied as well as approaches to prevent the free radical mediated cell damage which occurs.